Summary of Work: The lab is studying the molecular mechanisms whereby various G-protein-coupled receptors modulate the functional properties of voltage-gated calcium channels (VGCC) in rat sympathetic neurons and mouse pancreatic B-cells. By understanding the regulation of these channels in sympathetic neurons, we may understand better 1) how blood pressure is controlled, and 2) how presynaptic N-type VGCC control the release of neurotransmitter in the brain. The regulation of VGCC in pancreatic B-cells will help to understand the regulation of insulin secretion by these cells. Many types of receptor/G protein-coupled pathways have been shown to regulate VGCC, although the regulation of these pathways by Ca2+-dependent processes is largely unknown. To study this regulation, we have utilized patch-clamp recording technqiues in acutely dissociated cells. Our efforts have focused primarily on two of these G-protein-coupled receptor pathways; a pertussis toxin (PTX)- sensitive G-protein pathway (activated by nonadrenergic a2 and somatostatin receptors) that inhibits the N-type VGCC in rat sympathetic neurons, and a non-PTX-sensitive G-protein pathway (possibly G1 or G11) that is activated via the muscarinic subtype of acetylcholine (ACh) receptor that inhibits both N-type and L-type VGCC in the rat sympathetic neurons and the L-type channels in mouse pancreatic B-cells. We have further elucidated the Ca2+-dependent regulation of these two pathways. Our most notable discoveries over the past 12 months are 2-fold. First, we have discovered that for the PTX-sensitive pathway, if the Ca2+- calmodulin regulated protein phosphatase calcineurin is inhibited, the ability of this PTX-sensitive G-protein pathway to inhibit the N-type VGCC decreases by 50%. This effect could help to explain why the immunosuppressant drugs and certain classes of pesticides (which are potent inhibitors of calcineurin) induce hypertension in the peripheral nervous system and neurotoxic damage in the brain. Our second notable discovery relates to the regulation of VGCC by muscarinic receptor activation in pancreatic B-cells. Although we have further studied the regulation of this pathway in sympathetic neurons, our discovery that this pathway regulates the L-type VGCC in pancreatic B-cells helps to understand at a molecular level the regulation of insulin secretion in these cells.